Optimal method for isolation of human peritoneal mesothelial cells from clinical samples of omentum

INTRODUCTION: Human peritoneal mesothelial cells (HPMC) are a valuable research tool for understanding the molecular biology of several pathologies, in both monolayer and three dimensional models. We compared different methods of HPMC isolation and assessed their outcome as well as fibroblast contamination, a common problem encountered during isolation. METHODS: 1-3cm(3) samples of omentum were collected from 40 consenting patients undergoing elective gastrointestinal surgery. A total of 11 samples were incubated in 0.05% trypsin solution for 20 minutes at 37 degrees C (group A) and 29 in 0.25% trypsin (15 samples for 10 minutes (group B) and 14 for 20 minutes (group C)). Following digestion cells were re-suspended and cultured in supplemented Ham's F-12 medium containing 10% foetal calf serum (FCS), penicillin-streptomycin, glutamine, insulin, transferrin and hydrocortisone. Positive outcomes were absence of fibroblast contamination and satisfactory HPMC growth to confluence in a characteristic cobblestone pattern. Cytokeratins 5, 8, 18, Vimentin, Ber-Ep4 and Factor VIII were used to characterise HPMC and fibroblasts by immunohistochemistry. RESULTS: None of the 11 samples in group A yielded HPMC. 14 of 29 samples digested with 0.25% trypsin yielded HPMC: 10 of 14 yielded HPMC in group C versus four of 15 samples in group B (p = 0.02). Fibroblast contamination occurred in eight samples in group B versus three in group C. CONCLUSION: Optimal results are achieved with a 20 minute digestion in 0.25% trypsin. Fibroblast contamination could not be avoided completely. Other factors may minimise fibroblast contamination such as minimal tissue manipulation and early collection during surgery.     

Voiding urosonography: the study of the urethra is no longer a limitation of the technique

Background  Voiding urosonography (VUS) has proved to be a reliable method for the study of vesicoureteric reflux (VUR). Early reports considered it inadequate for imaging the male urethra. Objective  To determine the usefulness of contrast-enhanced VUS for the study of the urethra. Material and methods  A total of 208 children aged 2 days to 10 years underwent VUS to confirm or exclude VUR for different reasons (n = 150) or for follow-up (n = 58). Patients with unconfirmed suspicion of VUR (99 boys and 51 girls) also underwent VUS for the study of the urethra. Examinations were performed using a harmonic imaging mode specific for contrast (Levovist) enhancement. We used a 6–4-MHz convex probe and a transperineal and/or a transpelvic approach. Results  The neck of the bladder and the entire urethra were visualized in all patients (n = 150). The male urethra was considered normal in 95 boys (95.95%). We diagnosed posterior urethral valves in two patients, diverticulum of the prostatic utricle in one, and diverticulum of the anterior urethra in one. All abnormal cases were confirmed using conventional voiding cystourethrography. Conclusion  VUS can replace voiding cystourethrography as the method of choice for the initial study of suspected VUR in children. This work was presented at the 44th Annual Meeting of the European Society of Paediatric Radiology, Barcelona, Spain, 3–7 June 2007.     

Neoadjuvant continuous infusion of weekly 5-fluorouracil and escalating doses of oxaliplatin plus concurrent radiation in locally advanced oesophageal squamous cell carcinoma: results of a phase I/II trial

Oxaliplatin and 5-fluorouracil have a significant activity in locally advanced oesophageal squamous cell cancer (OSCC). However, their optimal dosage and efficacy when combined with concurrent radiotherapy as neoadjuvant treatment are unknown. This non-randomised, phase I/II study aimed to define the maximum tolerated dose (MTD) and assessed the histopathological tumour response rate to neoadjuvant oxaliplatin in weekly escalating doses (40, 45, 50 mg m(-2)) and continuous infusional 5-fluorouracil (CI-5FU; 225 mg m(-2)) plus concurrent radiotherapy. Patients had resectable OSCC. Resection was scheduled for 4-6 weeks after chemoradiotherapy. During phase I (dose escalation; n=19), weekly oxaliplatin 45 mg m(-2) plus CI-5FU 225 mg m(-2) was established as the MTD and was the recommended dosage for phase II. Oesophageal mucositis was the dose-limiting toxicity at higher doses. During phase II, histopathological responses (<10% residual tumour cells within the specimen) were observed in 10 of 16 patients (63%; 95% confidence interval: 39-82%). Overall, 16 of the 25 patients (64%) who underwent resection had a histopathological response; tumour-free resection (R0) was achieved in 80%. Neoadjuvant weekly oxaliplatin 45 mg m(-2) plus CI-5FU 225 mg m(-2) with concurrent radiotherapy provides promising histological response rates and R0 resection rates in locally advanced OSCC.     

Nonlinear local electrovascular coupling. II: From data to neuronal masses

In the companion article a local electrovascular coupling (LEVC) model was proposed to explain the continuous dynamics of electrical and vascular states within a cortical unit. These states produce certain mesoscopic reflections whose discrete time series can be reconstructed from electroencephalography (EEG) and functional magnetic resonance imaging (fMRI). In this article we develop a recursive optimization algorithm based on the local linearization (LL) filter and an innovation method to make statistical inferences about the LEVC model from both EEG and fMRI data, i.e., to estimate the unobserved states and the unknown parameters of the model. For a better understanding, the LL filter is described from a Bayesian point of view, providing the particulars for the case of hybrid data (e.g., EEG and fMRI), which could be sampled at different rates. The dynamics of the exogenous synaptic inputs going into the cortical unit are also estimated by introducing a set of Gaussian radial basis functions. In order to study the dynamics of the electrical and vascular states in the striate cortex of humans as well as their local interrelationships, we applied this algorithm to EEG and fMRI recordings obtained concurrently from two subjects while passively observing a radial checkerboard with a white/black pattern reversal. The EEG and fMRI data from the first subject was used to estimate the electrical/vascular states and parameters of the LEVC model in V1 for a 4.0 Hz reversion frequency. We used the EEG data from the second subject to investigate the changes in the dynamics of the electrical states when the frequency of reversion is varied from 0.5-4.0 Hz. Then we made use of the estimated electrical states to predict the effects on the vasculature that such variations produce.     

Predictors of in-hospital mortality present at admission among patients hospitalised because of decompensated heart failure

Chronic heart failure (HF) is associated with a poor prognosis and causes considerable mortality. The aim of this study was to identify the admission characteristics useful to predict in-hospital mortality in patients admitted because of decompensation of HF. We evaluated 414 patients (age 76.2 years, 57% women). The hospital mortality rate was 11.1%. We identified 4 independent predictors of mortality: low Barthel index (odds ratio 1.03; 95% confidence interval 1.01-1.04), creatinine level >200 mumol/l (odds ratio 3.40; 95% confidence interval 1.51-7.66), peripheral oedema (odds ratio 3.12; 95% confidence interval 1.28-7.58) and the protective effect of the new onset of the disease (odds ratio 0.2; 95% confidence interval 0.08-0.77). In conclusion, the mortality of patients admitted to the hospital with an exacerbation of HF can be predicted if either poor functional capacity, renal insufficiency, peripheral oedema or previous diagnoses of HF are present. This clinical finding may help clinicians in their decision making in HF in the emergency room.     

Immune response to vaccination against Argentine hemorrhagic Fever in an area where different arenaviruses coexist

Neutralizing antibody (NT Ab) titers to Candid #1 (C#1) vaccine against Argentine hemorrhagic fever were studied for 2 years post-vaccination in 330 volunteers, to assess whether the kinetics and/or magnitude of this immune response is modified by previous infection with the arena viruses Junin (JUN) and lymphocytic choriomeningitis (LCM). A total of 160 volunteers received C#1, distributed as follows: without detectable pre-infection with arenaviruses (n = 54); with pre-existing antibodies to JUN (n = 55); with pre-existing antibodies to LCM (n = 51). The remaining 170 individuals received placebo. Levels of anti-JUN NT Ab displayed a trend in which titers increased with the virulence of the infecting strain, from C#1 (X = 49), through subclinical JUN infection (X = 229), vaccination following subclinical infection (X = 367) to JUN clinical infection (X =773). It was also found that the mean titer of NT Ab to C#1 did not vary significantly during 2 years of study and was: a) significantly lower than that elicited by wild strains of JUN, both clinical and subclinical infections (p < 0.01); b) significantly increased the titers of pre-existing anti-JUN Ab (p < 0.01); and c) was not modified by pre-existing anti-LCM Ab (p > 0.05). These data indicate that the immune response to C#1 boosts pre-existing immunity to JUN virus and is not changed by previous experience with LCM virus.